Production of ethanol by microorganisms provides an alternative energy source to fossil fuels and is therefore an important area of current research. It is desirable that microorganisms producing ethanol, as well as other useful products, be capable of using xylose as a carbon source since xylose is the major pentose in hydrolyzed lignocellulosic materials, and therefore can provide an abundantly available, low cost carbon substrate. Zymomonas mobilis and other bacterial ethanologens which do not naturally utilize xylose may be genetically engineered for xylose utilization by introduction of genes encoding 1) xylose isomerase, which catalyses the conversion of xylose to xylulose; 2) xylulokinase, which phosphorylates xylulose to form xylulose 5-phosphate; 3) transketolase; and 4) transaldolase.
There has been success in engineering Z. mobilis strains for xylose metabolism (U.S. Pat. No. 5,514,583, U.S. Pat. No. 5,712,133, U.S. Pat. No. 6,566,107, WO 95/28476, Feldmann et al. (1992) Appl Microbiol Biotechnol 38: 354-361, Zhang et al. (1995) Science 267:240-243), as well as a Zymobacter palmae strain (Yanase et al. (2007) Appl. Environ. Microbiol. 73:2592-2599). However, typically the engineered strains do not grow and produce ethanol as well on xylose as on glucose. For this engineering, genes encoding the heterologous proteins for xylose metabolism have been expressed from promoters that are active in Z. mobilis cells, typically the promoter of the Z. mobilis glyceraldehyde-3-phosphate dehydrogenase gene or the promoter of the Z. mobilis enolase gene. Strains engineered for xylose utilization have been adapted by serial passage on xylose medium, resulting in strains with improved xylose utilization as described in U.S. Pat. No. 7,223,575 and commonly owned and co-pending U.S. Patent App. Publication No. US20080286870. However the genetic basis for the improvement had not been determined.
There remains a need for genetically engineered strains of Zymomonas, and other bacterial ethanolagens, having improved xylose utilization. Applicants have discovered mutant promoters having increased activity that can be used for expressing xylose utilization genes, which activity confers to engineered strains comprising these promoters improved xylose utilization. The promoters may be used for expression of other genes.